Table 4.
Primers and corresponding assays for diagnosis of Colletotrichum spp. On soybean.
| Target Gene | Target Species (Specificity) | Primer/Probe (Combination) | Sequence (5′-3′) | Tm (°C) | Fragment Length (bp) | Assay | Ref. |
|---|---|---|---|---|---|---|---|
| cox1 | C. chlorophyti | cox1AF | CCTGGTATAAGATTACATAAG | 55 | 115 | qPCR a | [65] |
| cox1AR | CTGTAAGTACCATAGTAATTG | ||||||
| cox1 | C. sojae | cox18AF | ACATTTATCAGGAGTAAGTAG | 55 | 77 | qPCR a | [65] |
| cox18AR | TTCCAGGTGTTCTCATAT | ||||||
| cox1 | C. incanum | cox6AF-2 | ATGAACATTATATCCTCCTT | 55 | 115 | qPCR a | [65] |
| cox6AR-2 | ATTAACTGCTCCTAATAAAC | ||||||
| cox1 | C. truncatum | cox15BF | TTATGCCAGCCTTAATAG | 55 | 117 | qPCR a | [65] |
| cox15BR | AAGATGGTGGTAATAATCA | ||||||
| ITS | C. gloeosporioides | Colg 1 | AACCCTTTGTGAACATACC | 63 | 443 | qPCR | [62] |
| Colg 2 | CCCTCCGGATCCCAG | ||||||
| ITS | C. truncatum | Colg 1 | AACCCTTTGTGAACATACC | 63 | 375 | qPCR a | [62] |
| CT 2 | CTTTAAGGGCCTACGTCAA | ||||||
| ITS | C. acutatum | CaITS_F701 | GGATCATTACTGAGTTACCGC | 60 | 80 | qPCR | [66] |
| CaITS_R699 | GCCCGCGAGAGGCTTC | ||||||
| CaITS_R815 b | GCCCACGAGAGGCTTC | ||||||
| CaITS_P710 | TACCTAACCGTTGCTTCGGCGGG | ||||||
| ITS | C. acutatum | ACUT-F1 | CGGAGGAAACCAAACTCTATTTACA | 60 | 70 | qPCR | [67] |
| ACUT-R1 | CCAGAACCAAGAGATCCGTTG | ||||||
| ACUT-PB | CGTCTCTTCTGAGTGGCACAAGCA | ||||||
| ITS | C. gloeosporioides | GLOE-F1 | GGCGGGTAGGGTCYCCG | 60 | 101 | qPCR | [67] |
| GLOE-R2 | ACTCAGAAGAAACGTCGTTAAATCAG | ||||||
| GLOE-PB | CTCCCGGCCTCCCGCCYC | ||||||
| ITS | Colletotrichum sp. | COL GEN-F1 | TGCCTGTTCGAGCGTCATT | 60 | 111 | qPCR | [67] |
| COL GEN-R2 | CTACGCAAAGGAGGCTCCG | ||||||
| COL GEN-PB | AACCCTCAAGCWCYGCTTGGYKTTGG | ||||||
| IGS | C. lupini | CLF | CCCGAGAAGGCTCCAAGTA | 63 | PCR | [63] | |
| CLR | CATAAACGCCTAAGAACCGC | ||||||
| GAPDH | C. truncatum | ColT-F6 | TTGAGACCAAGTACGCTGTATGTATCAC | 60 | qPCR | [64] | |
| ColT-R5 | TTCTGCCTCACATCGAACTCTC | ||||||
| ColT-P | HEX-CAGCCTTCG/ZEN/ACTCTCGTTGGAAAA-IABkFQ | ||||||
| GS | C. gloeosporioides | F3 | GCTGCAGCCGGAAAATCC | 64 | LAMP | [72] | |
| B3 | GGCAGACTCGGAGAGACC | ||||||
| FIP (F1c + F2) | ACCGGCTCAGCTGCAACGC-ACACGAGCAAAAGGATACGC | ||||||
| BIP (B1c + B2) | TAATGCCTTTCACGACCTGCGG-CCGAGGCAATGATTCCTCAA | ||||||
| LF | CGGGCCAACGCTGGAAAA | ||||||
| LB | GGCGCAACAAAGCTGGG | ||||||
| Rpb1 | C. truncatum | F3 | ACGGAGAATACTCTCTGGGT | 62 | LAMP | [71] | |
| B3 | AGGATGTTGTGTGCCATCTC | ||||||
| FIP (F1c + F2) | GCCTTGTGTCGGACTCTGGG-GCAAGCTCCCGTTAACCA | ||||||
| BIP (B1c + B2) | ACAGCTTGTCGCCAAGTACGAG-GGGTGTGATCTGAGGCTCTT | ||||||
| LF | TGAATGTTGCCACAGCCGC |
a Eva Green based real-time PCR. These four primer pairs can be used together in two duplex reactions where the products are distinguished by their melting temperature. b Alternative reverse primer to cover intraspecific sequence variation.